WHO estimates that 75% of the cases of HIV infection worldwide occur as a result of sexual intercourse. Thus, development of a vaccine against HIV capable of conferring protective mucosal immunity, and especially local genital immunity, will be crucial for control of this epidemic. The objective of this project will be to identify an oral immunization regimen, using small animals (mice) as a model, which is likely to confer effective, long-lasting and protective mucosal immunity against HIV infection. This objective will be achieved through the following specific aims. 1) Determine an optimal oral adjuvant / HIV immunogen regimen. This question will be addressed by orally immunizing mice with combinations of three HIV subunit immunogens (gp120 protein, a peptide representing the primary neutralizing determinant on gp120 and tat protein) and three oral adjuvants (cholera toxin, iscoms and microspheres) in different doses and with different immunization regimens. Secretory antibody responses, by ELISA and ELISPOT, will be used to measure the mucosal immune response to these regimens. 2) Determine the ability of secreted antibodies to neutralize HIV. It will be important to determine whether the oral immunization regimens employed are capable of generating neutralizing antibodies to HIV. To determine this, secreted antibodies will be assayed in vitro for HIV virus neutralization (prevention of p24 expression and viral cytopathic effects) and inhibition of cell to cell virus transmission. 3) Determine whether cytotoxic responses (CTL and ADCC) to HIV can be generated in mucosal tissues after oral immunization. Cytotoxic T cells against HIV are likely to play a key role in protective mucosal immunity to HIV. Therefore, lymphocytes from mucosal tissues will be tested for CTL and ADCC activity using in vitro assays with HIV immunogen-expressing target cells. 4) Determine the ability of oral immunization to generate a local mucosal immune response to HIV. Local mucosal immune responses in the genital tract or rectum to HIV will be assessed by measuring antibodies to HIV (including neutralizing antibodies) in secretions from these sites. 5) Determine the duration of secretory antibody responses to HIV and the length of memory B and T cell responses. For an oral vaccine to be effective, antibodies in secretions must persist for long periods of time. Orally immunized mice will have secretions collected at various, intervals, up to 12 months after immunization, and assayed for antibodies to HIV. Memory responses will be assessed by collecting lymphocytes from mucosal tissues at various time points after oral immunization and assaying for in vitro responses to HIV antigens. 6) Determine whether oral immunization with HIV immunogens results in oral tolerance to these immunogens. This will be evaluated by systemically reimmunizing mice which have previously been orally immunized with the same HIV immunogen and evaluating the subsequent systemic immune response (ie. serum antibody response and cytotoxicity).